Analytical determinations and protein modifications will be performed in an effort to determine the nature of the mercaptan requirement for phosphoenol-pyruvate carboxykinase (PEPCK). Stereochemical requirements at the catalytic site will be investigated after synthesizing several substrate analogs and using kinetic and direct binding studies. The interaction of the substrates to the enzyme-Mn to the second power ion complex will be studied by high resolution 1H, 31P and 13C nuclear relaxation rate studies. The interaction of the substrate Cr(III) IDP to the enzyme will be studied by NMR and EPR techniques. The structure of the enolase-Mn-F-Pi complex which inhibits glycolysis will be further investigated by high field relaxation rate studies to obtain more definitive data concerning the structure of this complex. The environments of the F to the negative power and Pi ligands in their respective ternary and the quaternary complexes will be obtained by observing the chemical shifts and line widths of the bound ligands. This will, in part, be compared to studies performed with the substrates where environmental and kinetic data will be obtained. Yeast pyruvate kinase will be investigated in terms of the ligand binding and ligand structure in an effort to physically describe the nature of allosterism in this enzyme.